Well-defined diblock copolymers comprising thermoresponsive segments of poly(N-isopropylacrylamide-co-N,N-dimethylacrylamide) (P(IPAAm-co-DMAAm)) and hydrophobic segments of poly(d,l-lactide) were synthesized by combination of RAFT and ring-opening polymerization methods. Terminal conversion of thermoresponsive segments was achieved through reactions of maleimide or its Oregon Green 488 (OG) derivative with thiol groups exposed by cleavage of polymer terminal dithiobenzoate groups. Thermoresponsive micelles obtained from these polymers were approximately 25 nm when below the lower critical solution temperature (LCST) of 40 degrees C, and their sizes increased to an average of approximately 600 nm above the LCST due to aggregation of the micelles. Interestingly, the OG-labeled thermoresponsive micelles showed thermally regulated internalization to cultured endothelial cells, unlike linear thermoresponsive P(IPAAm-co-DMAAm) chains. While intracellular uptake of P(IPAAm-co-DMAAm) was extremely low at temperatures both below and above the micellar LCST, the thermoresponsive micelles showed time-dependent intracellular uptake above the LCST without exhibiting cytotoxicity. These results indicate that the new thermoresponsive micelle system may be a greatly promising intracellular drug delivery tool.

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A.K. and M.A. performed the experiments repeatedly and collected and analyzed the data. L.A. and S.W. prepared and provided the fibrinogen solutions in varying concentrations and provided technical inputs. S.A.K., M.A., and B.J. wrote the manuscript and prepared all the figures. S.C.A. and L.S. performed rheology and analyzed the resultant data. V.T. and M.C. facilitated the CM and CF cultures, cell coupling experiments, and their microscopic imaging along with images and writing of relevant sections in the manuscript. J.A. and Y.I. provided the furfuryl-gelatin and performed cytotoxicity assay for Rose Bengal. All authors reviewed the manuscript and provided their consent for publication. The manuscript was written through individual contributions of all authors. All authors have given approval to the final version of the manuscript.

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